Infection with simian type D retrovirus (SRV/D) is endemic in many populations of macaques. SRV/D is a significant pathogen in macaque colonies, being the etiologic agent of one form of simain acquired immune deficiency disease. In addition to its importance from an animal health perspective, SRV/D infection may also represent a significant confounding variable in the interpretation of research results obtained using macaques as experimental subjects. In recognition of the importance of establishing and maintaining breeding colonies of macaques free of SRV/D infection, the objects of this research are to develop enhanced serologic and virologic diagnostic tests for SRV/D infection and employ these improved assays in support of the NCRR-sponsored SPF rhesus macaque development program. We have successfully developed and validated a nested PCR which allows simultaneous screening for SRV/D serogroups 1, 2, and 3. PCR is more sensitive than either serologic testing or virus isolation for identification of SRV/D infected animals, capable of detection of from 1-10 viral genomic copies. Additionally, we are evaluating recombinant antigens expressed in yeast, bacterial, and baculovirus expression systems to improve both the sensitivity and specificity of serologic assays for SRV/D antibody. The nested PCR has been made available for screening SPF colony development, and to other facilities housing macaques used in NIH-funded biomedical research, and has proven a powerful tool in SRV/D control and elimination strategies. *KEY*SRV/D, Retrovirus, Macaque, Diagnosis, PCR, SPF